Properties of the cuticular proteins of Anopheles gambiae as revealed by serial extraction of adults.
Properties of the cuticular proteins of Anopheles gambiae as revealed by serial extraction of adults.
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DOI：
10.1371/journal.pone.0175423
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发表时间：
2017
期刊：
影响因子：
3.7
通讯作者：
Willis JH
中科院分区：
文献类型：
--
作者：
Zhou Y;Badgett MJ;Billard L;Bowen JH;Orlando R;Willis JH
角质层蛋白（CP）如何与甲壳素相互作用以及在角质层中相互作用仍然没有解决。我们采用LC-MS/MS鉴定了用PBS、EDTA、2- 8 M尿素和SDS连续提取后释放的5-6日龄冈比亚按蚊成虫的CP以及未提取的CP。将结果与已发表的转录本丰度时间、结构内和角质层内蛋白质的定位以及单个蛋白质的性质、长度、pI、组氨酸百分比、酪氨酸、谷氨酰胺和AAP[A/V/L]重复数的数据进行比较。13种蛋白质完全溶解，全部为CPR，大多数属于RR-1组。在可溶性组分和最终沉淀物中鉴定了11种CP，包括5种来自其他CP家族。仅从最终颗粒中检测到43个。这些包括CPRs和CPAP 1、CPF、CPFL、CPLCA、CPLCG、CPLCP和TWDL家族的成员，以及几个未分配到家族并命名为CPLX的低复杂度CP。对于给定的蛋白质，许多组氨酸或酪氨酸或谷氨酰胺似乎是交联的潜在参与者，因为我们不能鉴定出任何带有这些始终不存在的残基的肽。我们未能从任何CP的氨基末端回收肽。目前尚不清楚这是否意味着硬化中的该位置或阻止检测的某些修饰。可溶性CPRs的等电点低于最终沉淀中的那些，其他CP家族的大多数成员的等电点为8或更高。显然，除了分析不同的溶解度之外，还需要其他技术来了解CP之间以及与几丁质之间的相互作用。
How cuticular proteins (CPs) interact with chitin and with each other in the cuticle remains unresolved. We employed LC-MS/MS to identify CPs from 5–6 day-old adults of Anopheles gambiae released after serial extraction with PBS, EDTA, 2-8M urea, and SDS as well as those that remained unextracted. Results were compared to published data on time of transcript abundance, localization of proteins within structures and within the cuticle, as well as properties of individual proteins, length, pI, percent histidine, tyrosine, glutamine, and number of AAP[A/V/L] repeats. Thirteen proteins were solubilized completely, all were CPRs, most belonging to the RR-1 group. Eleven CPs were identified in both soluble fractions and the final pellet, including 5 from other CP families. Forty-three were only detected from the final pellet. These included CPRs and members of the CPAP1, CPF, CPFL, CPLCA, CPLCG, CPLCP, and TWDL families, as well as several low complexity CPs, not assigned to families and named CPLX. For a given protein, many histidines or tyrosines or glutamines appear to be potential participants in cross-linking since we could not identify any peptide bearing these residues that was consistently absent. We failed to recover peptides from the amino-terminus of any CP. Whether this implicates that location in sclerotization or some modification that prevents detection is not known. Soluble CPRs had lower isoelectric points than those that remained in the final pellet; most members of other CP families had isoelectric points of 8 or higher. Obviously, techniques beyond analysis of differential solubility will be needed to learn how CPs interact with each other and with chitin.